Anexelekto (also referred to as “AXL”, “UFO”, “ARK”, or “TYRO7”; hereinafter referred to as “AXL”) is a receptor tyrosine kinase that exists on the cell membrane (Non-patent Document 1). It is said to be responsible for signal transduction to downstream molecules through its autophosphorylation, which occurs after it binds to the ligand Gas6 (growth arrest specific gene 6) (Non-patent Document 2).
AXL is presumed to have molecular functions involved in cell growth enhancement, suppression of apoptosis, cell migration, and cell adhesion. Experimentally observed phenomena in cells treated with Gas6 protein support this presumption. Reported experimental results include suppression of cell death and enhancement of cell growth in rat vascular smooth muscle (Non-patent Documents 3 and 4), acceleration of cell growth and the suppression of cell death after serum starvation in mouse NIH3T3 cells (Non-patent Documents 5 and 6), promotion of cell growth in mouse cardiac fibroblasts (Non-patent Document 7), enhancement of cell growth in human prostate cancer cells (Non-patent Document 8), enhancement of cell growth and infiltration and suppression of cell death in human gastric carcinoma cells (Non-patent Document 9), enhancement of the migration ability of human and rat vascular smooth muscle cells (Non-patent Document 10), enhancement of the cell migration ability of mouse neurons (Non-patent Document 11), and aggregation of cells highly expressing mouse AXL (Non-patent Document 12).
Similarly, PI3K-Akt pathway and MAPK pathway are said to function as downstream pathways of the signal transduction mediated by AXL based on molecular analyses of intracellular signals after treatment with Gas6 (Non-patent Document 2). An analysis with a yeast two-hybrid method using an AXL intracellular region as bait confirmed direct molecular interaction with these downstream pathways. As a result, GrbB2/PI3K/p55γ/SOCS-1/NcK2/RanBP2/C1-TEN were identified (Non-patent Document 13). The interactions of these molecules suggest the presence of intracellular signal transduction pathways as downstream from the AXL signal. Furthermore, the observed interactions support the presumption that AXL functions in cell growth enhancement, suppression of apoptosis, cell migration, and cell adhesion. AXL has also been identified as a gene highly expressed when TNFα-induced cell death of mouse fibroblasts is suppressed by IL-15. The suppression of TNFα-induced cell death was abolished by suppressing AXL expression, and the phosphorylation of IL-15 receptors and AXL was enhanced by treatment with IL-15. These experimental findings also suggest that signal transduction is mediated by the complex of AXL and IL-15 receptors (Non-patent Document 14).
Tumorigenicity of nude mice has been reported to disappear as a result of inhibiting Gas6-dependent phosphorylation of AXL in human glioma lines overexpressing the AXL dominant negative form (Non-patent Document 15). However, there were no reports or suggestions of anti-AXL antibody that inhibits phosphorylation and its existence remained unclear.
AXL is a single-pass transmembrane receptor tyrosine kinase, and the extracellular region is composed of two immunoglobulin-like domains (referred to as IgD1 and IgD2) and two fibronectin type III domains (referred to as FND1 and FND2) from the N-terminal side (Non-patent Document 1). Although FND is known to be expressed in molecules such as neural cell adhesion molecules and nephrins involved in intercellular adhesion, detailed functions of FND in AXL are unclear (Non-patent Document 16).
AXL has been identified as an oncogene that retains an inherent ability to transform cells, and has been studied as a carcinogenesis-related molecule. Many analyses of AXL expression have been reported on the protein and mRNA. The high expression of AXL protein has been reported in human cancer tissues and cancer cells, including lung cancer (Non-patent Document 17), breast cancer (Non-patent Document 18), ovarian cancer (Non-patent Document 19), thyroid cancer (Non-patent Document 20), melanoma (Non-patent Document 20), renal cancer (Non-patent Document 21), gastric cancer (Non-patent Document 9), and glioma (Non-patent Document 22). Furthermore, the high expression of AXL protein is suggested by the high level of AXL mRNA in esophageal cancer (Non-patent Document 23), colon cancer (Non-patent Document 24), and acute myeloid leukemia (Non-patent Document 25). There are also reports of inhibition of lumen formation via suppression of AXL by RNAi in HUVEC (Non-patent Document 26), reduced tumor-forming ability of human breast cancer cells in mice resulting from constitutive suppression of AXL (Non-patent Document 26), and reduced tumor-forming ability of human glioma cells in mice resulting from a constitutive high expression of dominant negative mutants (Non-patent Document 22). The involvement of AXL molecular functions in tumor growth is strongly suggested.
Polyclonal antibodies to the extracellular domain of AXL have been reported to have a migration inhibitory action on highly invasive breast cancer cell lines (Patent Document 1). However, non-clinical studies showed that drugs demonstrating cancer-cell-migration-inhibitory action do not necessarily demonstrate antitumor activity. For example, matrix metalloproteinase (hereinafter abbreviated to “MMP”) has been known to promote tumor infiltration and migration. Thus, as candidates of anticancer agents, attention has been focused on various matrix metalloproteinase inhibitors that inhibit the enzyme activity of MMP, and clinical studies have been conducted on various pharmaceutical agents such as Batimastat, Marimastat, and Prinomastat. However, antitumor effects have not been observed in clinical trials (Non-patent Document 27).
Accordingly, there have been no reports or suggestions and it remains unknown whether antibodies that bind to a specific region of AXL have antitumor effects particularly in vivo, whether they can reduce AXL expression levels, and whether they can suppress cancer.